The overarching goal of this Program is to determine the vulnerabilities of the HIV-1 Env protein cell entry machine as a target for disease intervention by identifying Env inhibitors, defining their structural mechanisms of action, and using a structure-mechanism framework as a guide to optimize antagonist functions. Inhibition of the initial entry of HIV-1 into host cells remains a compelling, yet elusive means to prevent infection and spread of the virus. Inhibitors of HIV-1 Env that can either block cell interactions, inactivate the trimeric virus spike protein complex before receptor encounter or disrupt receptor-induced conformational changes in the Env would hold great promise of inhibiting initial HIV-1 infection. Such inhibitors would provide virus-targeted molecular weapons both to prevent AIDS transmission, a global health priority, and to treat already-infected individuals. In spite of the great potential of Env inhibitors for AIDS intervention, structural complexity and polymorphisms of the Env proteins have presented significant challenges to progress. Nonetheless, the efforts of our Program have led to the development of two classes of Env gp120 inhibitors that utilize the highly conserved CD4 binding site, but with very different modes of action. Investigation of these inhibitors has defined unique pathways to engage the virus Env trimer and cause both inactivation of the virus and blockade of virus entry into the host cell. Our Program is ideally positioned to take advantage of these new results through state-of-the-art structure- and mechanism-based approaches, achieved by the collaborative nature of our multi-institutional research team, with strong expertise in high-resolution structure determination, structural dynamics, kinetic, thermodynamic and structural mechanisms of protein-protein interactions, chemical design and synthesis, computational methods, and virology. We will apply this team approach to structure-based design and mechanistic investigations of inhibitor chemotypes that we have already developed, and new inhibitor chemotypes as they are discovered in our own and other laboratories. Overall, the Program will provide a broad-based research infrastructure to identify new paths for the discovery of preventive and therapeutic agents that block HIV-1 Env function.